Generation of cell-type-specific proteomes of neurodevelopment from human cerebral organoids

Sofia Melliou; Phedias Diamandis

doi:10.1016/j.xpro.2022.101774

Generation of cell-type-specific proteomes of neurodevelopment from human cerebral organoids

STAR Protoc. 2022 Oct 21;3(4):101774. doi: 10.1016/j.xpro.2022.101774. eCollection 2022 Dec 16.

Authors

Sofia Melliou¹, Phedias Diamandis²

Affiliations

¹ Princess Margaret Cancer Centre, 101 College Street, Toronto, ON M5G 1L7, Canada; Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON M5S 1A8, Canada. Electronic address: sophia.melliou@mail.utoronto.ca.
² Princess Margaret Cancer Centre, 101 College Street, Toronto, ON M5G 1L7, Canada; Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON M5S 1A8, Canada; Laboratory Medicine Program, University Health Network, 200 Elizabeth Street, Toronto, ON M5G 2C4, Canada; Department of Medical Biophysics, University of Toronto, Toronto, ON M5S 1A8, Canada. Electronic address: p.diamandis@mail.utoronto.ca.

Abstract

Characterization of cerebral organoids has been challenging due to their heterogeneous nature. Here, we optimized a protocol to streamline the generation of FACS-purified cell populations from human cerebral organoids for proteomic analysis with liquid chromatography tandem mass spectrometry (LC-MS/MS). We describe the procedures for enzymatic dissociation of organoids into single-cell suspension, the generation of cell-type-specific lysates, peptide extraction, and proteomic analysis. This generalizable approach can be used to study temporal and cell-type-specific protein dynamics in developing cerebral organoids. For complete details on the use and execution of this protocol, please refer to Melliou et al. (2022).

Keywords: Cell isolation; Flow cytometry/Mass cytometry; Mass spectrometry; Neuroscience; Organoids; Proteomics.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Chromatography, Liquid / methods
Humans
Organoids
Proteome*
Proteomics* / methods
Tandem Mass Spectrometry / methods

Substances

Proteome

Grants and funding

CIHR/Canada